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Gene expression profiling of single cells from archival tissue with laser-capture microdissection and Smart-3SEQ.


ABSTRACT: RNA sequencing (RNA-seq) is a sensitive and accurate method for quantifying gene expression. Small samples or those whose RNA is degraded, such as formalin-fixed paraffin-embedded (FFPE) tissue, remain challenging to study with nonspecialized RNA-seq protocols. Here, we present a new method, Smart-3SEQ, that accurately quantifies transcript abundance even with small amounts of total RNA and effectively characterizes small samples extracted by laser-capture microdissection (LCM) from FFPE tissue. We also obtain distinct biological profiles from FFPE single cells, which have been impossible to study with previous RNA-seq protocols, and we use these data to identify possible new macrophage phenotypes associated with the tumor microenvironment. We propose Smart-3SEQ as a highly cost-effective method to enable large gene expression profiling experiments unconstrained by sample size and tissue availability. In particular, Smart-3SEQ's compatibility with FFPE tissue unlocks an enormous number of archived clinical samples; combined with LCM it allows unprecedented studies of small cell populations and single cells isolated by their in situ context.

SUBMITTER: Foley JW 

PROVIDER: S-EPMC6836736 | biostudies-literature | 2019 Nov

REPOSITORIES: biostudies-literature

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Gene expression profiling of single cells from archival tissue with laser-capture microdissection and Smart-3SEQ.

Foley Joseph W JW   Zhu Chunfang C   Jolivet Philippe P   Zhu Shirley X SX   Lu Peipei P   Meaney Michael J MJ   West Robert B RB  

Genome research 20190913 11


RNA sequencing (RNA-seq) is a sensitive and accurate method for quantifying gene expression. Small samples or those whose RNA is degraded, such as formalin-fixed paraffin-embedded (FFPE) tissue, remain challenging to study with nonspecialized RNA-seq protocols. Here, we present a new method, Smart-3SEQ, that accurately quantifies transcript abundance even with small amounts of total RNA and effectively characterizes small samples extracted by laser-capture microdissection (LCM) from FFPE tissue.  ...[more]

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